Abstract Recentadvancesinsingle?celltranscriptomicsindissociatedcellshavepermittedtheunbiasedclassification ofunique,molecularly?definedcelltypesinmanybrainregions.However,linkingthesemolecularly?defined celltypestotheircorrespondingmorphological,physiological,andfunctionalphenotypesremainsamajorchal? lengeinthefield.Wehaveassembledaninterdisciplinaryteamcapableofcombiningandfurtheroptimizing cutting?edgetechnologiesincludingPatch?seq(amethodwedevelopedthatcombineswhole?cellpatch?clamp recordingsandsingle?cellRNAsequencing),multi?photoncalciumimaging,multiplexedfluorescentinsituhy? bridization(MERFISH),andstate?of?artmachinelearningtoaddressthisgapinknowledge.Inaim1,duringthe firsttwoyears,wewillprovideacomprehensivecensusofthecelltypesthatcomprisethemouseprimaryvisual cortexcircuitrybylinkingsinglecellRNA?seq,morphological,andinvitroelectrophysiologicaldata.Duringthe last three years, we will define the cell types of higher order visual areas. In addition, we will characterize a distributednetworkoffivesubcorticalregionsinvolvedinthecontrolofinnatesocialbehaviors,includingthe ventromedialhypothalamicnucleus,medialpreopticnucleus,anteriorhypothalamicnucleus,posteriorbednu? cleus of the stria terminalis, and posterior medial amygdala. By studying both cortical and subcortical brain regions,wewillbeabletocomparesimilaritiesanddifferencesacrossthoseregions,aswellascompareprinci? plesofcelltypeorganizationbetweenevolutionarilyancientsubcorticalandmorerecentlyevolvedcorticalre? gionsofthebrain.Inaim2,wewillstudythefunctionalpropertiesoftranscriptomically?definedcelltypesin thevisualcortexofthemouse(areasV1,LM,PMandAM).Wewillperformmulti?photoncalciumimagingin behavingmiceduringthepresentationofavarietyofvisualstimulitocharacterizeindetailthereceptivefield propertiesoftheseneuronsfollowedbyMERFISHtoidentifythegeneticprofileoftherecordedneurons.We willemploythistobothCrelinesthatlabelknownbroadclassesofneuronsaswellasdenseimagingofcortical populationstoprovideacomplete,specific(i.e.inthesameanimal)characterizationofbothcelltypefunction andmolecularprofiling.Thecombineddatafromthesetwospecificaimspromisetoprovidethemostcomplete understandingofcelltypestodate,includingexpressionprofiles(e.g.ionchannelandreceptorlevels)morphol? ogy,single?cellelectrophysiology,andinvivofunctionalproperties.Themethodsandpipelinesthatwillbeop? timizedinthissectionoftheproposalwillalsolaythefoundationtofurtherapplythesemethodsindifferent partsofthebrainaswellasstudyanimalmodelsofdiseases.